Sequences from human immunodeficiency virus (HIV)-1 isolates derived form three individuals over time periods ranging from four to nine years were analyzed phylogenetically. The env gene sequences were found to acquire diversity at a faster rate than the nef gene sequences. As a result, nef sequences but not env sequences, from the same individual or cluster from different times were monophyletic. Since the difference between inter- and intra-patient variability is maintained over a longer period of time, nef may be of greater use than env in identification of infection clusters. Both the env and nef sequences of a laboratory worker infected with HIV-1 by occupational exposure acquired diversity at a slower rate than the sequences of viruses isolated from two male homosexuals. This may be due to the lesser complexity of the infecting virus.